Characterization of MyDNMT1 and Changes of Global DNA Methylation during the Embryonic Development in Mizuhopecten yessoensis

DNA methylation is a critical epigenetic mechanism that influences gene transcription, genomic stability, X-chromosome inactivation and other factors, and appropriate DNA methylation is crucial in development. DNA methyltransferase 1 （DNMT1） plays an important role in maintaining the established methylation pattern during DNA replication. Although the effect of DNA methylation on embryonic development has been well known in vertebrates, little research has been carried out in invertebrates, especially in marine bivalves. In this study, the DNMT1 gene （MyDNMT1） was firstly identified from Mizuhopecten yessoensis. The full-length cDNA of MyDNMT1 was 5 039 bp, consisted of a 5＇ untranslated region （5＇-UTR） of 79 bp, a 3＇ untranslated region （3＇-UTR） of 199 bp, and a 4 761 bp open reading frame （ORF） encoding a peptide of 1 586 amino acids without a putative signal peptide. The relative mRNA expression level of MyDNMT1 was measured during the embryonic development of M. ydssoensis using real-time PCR, which revealed that the level at stage zygote and trochophore were significantly higher than that at other stages. We further examined the global DNA methylation during development by colorimetric method. The results showed that the methylation level was increased and reached the peak at blastula stage, then dramatically decreased, and fluctuated at early D-shaped larva stage. This study provided greater insight into the DNA methylation of embryonic development, which obtained a better understanding of the relationship between the DNA methylation and the embryonic development in bivalve mollusks.

Chronic cerebrovascular hypoperfusion affects global DNA methylation and histone acetylation in rat brain

DNA methylation and histone acetylation can be modified by various pathological or physiological factors such as hypoxia,thus influencing gene expression.In this study,we investigated the changes of global DNA methylation and histone acetylation and the related enzymes in rat brain after chronic cerebrovascular hypoperfusion by bilateral common carotid occlusion（2-VO） surgery.Colorimetric and immunohistochemistry staining were used to evaluate the global DNA methylation and histone acetylation levels,respectively.The expressions of DNA methyltransferase 1/3a（DNMT1/3a）,methyl-CpG binding domain protein 2（MBD2）,histone deacetylase 3（HDAC3） and acetyltransferase（HAT） were assessed by Western blot.We found that the level of global DNA methylation was decreased to 31.7%（P ＆lt;0.01） of the sham-operated group at 10 days and increased by 30%（P ＆lt;0.01） compared with the sham group at 90 days after 2-VO surgery.DNMT3a expression was down-regulated to 75.7% of the sham group,while MBD2 expression was up-regulated by 95% compared with sham group at 90 days after 2-VO.The histone H3 acetylation level was markedly decreased to 75.3% of the sham group at 10 days and 73.5% at 90 days after 2-VO,while no significant change was found for histone H4 acetylation.HDAC3 expression was markedly down-regulated to 36% of the sham group,whereas cAMP-response element binding protein expression was up-regulated by 33.6% compared with the sham group at 90 days after 2-VO.These results suggest that chronic cerebrovascular hypoperfusion influences global DNA methylation and histone acetylation levels through the related enzymes,and therefore might contribute to several neurodegenerative diseases.

Global methylation status of sperm DNA in carriers of chromosome structural aberrations

Male infertility might be clearly associated with aberrant DNA methylation patterns in human spermatozoa. An association between oxidative stress and the global methylation status of the sperm genome has also been suggested. The aim of the present study was to determine whether the global sperm DNA methylation status was affected in the spermatozoa of carriers of chromosome structural aberrations. The relationships between the 5-methylcytosine （msC） levels in spermatozoa and chromatin integrity status were evaluated. The study patients comprised male carriers of chromosome structural aberrations with reproductive failure （n = 24）, and the controls comprised normozoospermic sperm volunteers （n = 23）. The global msC level was measured using thin-layer chromatography （TLC） and immunofluorescence （IF） techniques. The sperm chromatin integrity was assessed using aniline blue （AB） staining and TUNEL assay. The mean msC levels were similar between the investigated chromosome structural aberrations carriers （P） and controls （K）. However, sperm chromatin integrity tests revealed significantly higher values in chromosomal rearrangement carriers than in controls （P 〈 0.05）. Although the potential relationship between sperm chromatin integrity status and sperm DNA fragmentation and the msC level juxtaposed in both analyzed groups （P vs K） was represented in a clearly opposite manner, the low chromatin integrity might be associated with the high hypomethylation status of the sperm DNA observed in carriers of chromosome structural aberrations.